Chronic myelomonocytic leukemia (CMML) is a devastating cancer for which there is currently no effective therapy. Approximately 20% of CMML cases evolve to acute myelogenous leukemia (AML) soon after their initial diagnosis. Oncogenic NRAS mutations, which are among the most frequently identified genetic mutations in myeloid diseases, are identified in 17-60% of CMML cases, including cases that transform to AML. However, it remains elusive how oncogenic, endogenously arising NRAS mutations leads to CMML and its transformation to AML. Recently, we established a mouse bone marrow transplantation model harboring an oncogenic G12D mutation in the endogenous Nras locus in which ~95% of recipient mice develop a myeloproliferative (MP) disease remarkably resembling the MP variant of human CMML. Our preliminary results suggest that endogenous oncogenic Nras signaling promotes HSC proliferation and mobility rather than apoptosis and senescence. We propose that genetically altered HSCs initiate and maintain CMML in this model. In addition, similar to what occurs in patients with CMML, aberrant GM-CSF (granulocyte-macrophage colony stimulating factor) signaling is a signature of our model, primarily regulating expansion of granulocytic/monocytic precursors. We hypothesize that this aberrant signaling drives inappropriate cell growth and survival during disease initiation and progression, and thus could constitute a valuable therapeutic target. Because CMML occurs after a prolonged latency accompanied by multiple additional genetic lesions in our model, we further hypothesize that, as for human CMML, oncogenic NRAS cooperates with mutations in other genes to either induce CMML or lead to CMML transformation to AML. As a part of our long-term goal to understand the molecular and cellular mechanisms in tumor initiation, progression, and malignant transformation, in this application we propose: 1) To determine the effects of endogenous oncogenic Nras signaling on the properties of HSCs and examine whether HSCs expressing oncogenic Nras initiate and maintain CMML;2) To determine whether aberrant GM-CSF signaling is essential to establish and/or maintain oncogenic Nras-initiated CMML-like phenotypes;3) To identify novel pathogenic origins involved in CMML and/or its transformation to AML using CMML patient samples and to validate cooperating mutations of oncogenic NRAS in our murine model of CMML. Successful accomplishment of the proposed studies will not only provide insights into the pathogenesis, progression, and transformation of CMML, but may also lead to novel insights into HSC regulation, aberrant cytokine signaling, and cooperating mutations in oncogenic NRAS- associated myeloid diseases in general. PUBLIC HEALTH RELEVANCE: Chronic myelomonocytic leukemia (CMML) is a devastating disease that primarily affects the elderly. Although oncogenic NRAS mutations are frequently identified in CMML patients, its role remains unknown in initiation, progression, and malignant transformation of CMML. In this application, we propose to address this question using CMML patient samples and our new murine model of CMML. Furthermore, because oncogenic NRAS mutations are among the most frequently identified genetic mutations in myeloid diseases, successful completion of the proposed studies on HSC regulation, aberrant cytokine signaling, and validation of cooperating mutations of oncogenic NRAS have broad impact on understanding the role of NRAS mutations in hematological malignancies, including but not limiting to CMML.